1. Field of the Invention.
The present invention relates to processes for producing and purifying 2',3'-dideoxynucleosides, and to processes for producing 2',3'-dideoxy-2',3'-didehydronucleosides.
2. Discussion of the Background.
2',3'-Dideoxynucleosides and 2',3'-dideoxy-2',3'-didehydronucleosides display anti-viral activity but are prohibitively expensive to prepare industrially. For example, 2',3'-dideoxy-2',3'-didehydronucleosides of the formula (I): ##STR1## per se and 2',3'-dideoxynucleosides of the formula (II): ##STR2## which can be obtained by reducing 2',3'-dideoxy-2',3'-didehydronucleosides, both display anti-viral activity and can be utilized, for example, in the treatment of AIDS.
Didehydronucleosides can therefore be used either as drugs or as intermediates for the production of drugs useful to combat viruses (cf. Published Unexamined Japanese Patent Application No. 280500/86 and J. Med. Chem., 30, 440 (1977)). (The definition of substituent B is provided infra.)
Dideoxynucleosides, such as 2',3'-dideoxyadenosine [9-.beta.-D-(2',3'-dideoxyribofranosyl)adenine]; 2',3'-dideoxyinosine [9-.beta.- D-(2',3'-dideoxyribofranosyl)hypoxanthine]; and 2',3'-dideoxyguanosine [9-.beta.- D-([2',3'dideoxyribofranosyl)guanine], also possess powerful anti-viral activity. All of these materials are therefore expected to be useful as antiviral medicines, particularly as drugs for the treatment of AIDS which is an intractable disease of worldwide concern.
2',3'-Dideoxy-2',3'-didehydronucleosides can be produced by a method which uses ribonucleosides as raw materials (cf. J. Org. Chem., 39 (1974). Another method uses 2'-deoxyribonucleosides as raw materials (cf. J. Amer. Chem. Soc., 88, 1549 (1966) and J. Org. Chem., 32, 817 (1967)), etc.
The conventional method for producing 2',3'-dideoxynucleosides is by chemical deoxygenation of nucleosides at the 2 or 3 position, as described in Chem. Pharm. Bull., 22, 128 (1974). But reports on this method are very few however, and no industrial manufacturing process has yet been established on the basis of this basic method. The reasons for this are due to the facts that in this method (1) protective groups must be introduced prior to deoxygenation, (2) the reaction does not proceed smoothly because of the steric hindrance at the 2'- and 3'-positions, and (3) severe reaction conditions or powerful reagents cannot be used because nucelosides are unstable under such severe conditions. Nothing is so far known about a microbial process for producing 2',3'-dideoxynucleosides.
In all of these methods many steps are required and expensive materials must be used as the raw materials. The conventional chemical methods also have the problems that long reaction steps are involved and the product yield is low. These methods are consequently not advantageous from an industrial viewpoint. And hence, there has been a need for a new process for producing efficiently and inexpensively 2',3'-dideoxynucleosides in high yields.
There also has been a need for a method which can provide 2',3'-dideoxy-2',3'-didehydronucleosides industrially efficiently and inexpensively using readily available starting materials.
In an available process for producing 2',3'-dideoxyinosine (DDI), the oxygen atom at the 2'- or 3'-position of the nucleoside is eliminated (see Chem. Pharm. Bull., 22, 128 (1974)). However, this process has not been used widely because (1) protective groups must be introduced prior to the reaction, and (2) the deoxygenation reaction tends to be hindered by severe steric hindrance at the 2'- and 3'-positions.
In cases where DDI is produced from microbial or enzymatic action on a substrate such as 2',3'- dideoxyuridine (DDU) or 2,3-dideoxyribose-1-phosphoric acid, the reaction mixture obtained contains, in addition to the desired product (DDI), unreacted DDU, hypoxanthine (Hyp), uracil (Ura) formed by the decomposition of the substrate, and small quantities of nucleic acids formed as by-products.
Known purification treatments, such as concentration and recrystallization, are not suited for obtaining efficiently high purity DDI from these mixtures. This is because impurities, such as Ura, Hyp, etc., have solubilities lower than that of the desired DDI and, consequently, the crystals of DDI which as formed are contaminated with the impurities. This makes the purification of DDI exceedingly difficult.
In addition to this, DDI is susceptible to hydrolysis under either acidic or neutral conditions. It is therefore difficult to purify DDI by means of ion exchange treatment since an acid is utilized for this treatment and, DDI is consequently hydrolyzed into a 2,3-dideoxyribose residue and a hypoxanthine residue.
For the above reasons, the purification and isolation of DDI has been practiced only in laboratories by means of repeated liquid cyromatography or thin layer chromatography. No commercial process for the purification of DDI is available. There has therefore been a need for an industrially advantageous purification process which makes it possible to efficiently and inexpensively purify DDI.
Another process has been reported in which 2',3'-dideoxyadenosine (DDA) is subjected to enzymatic deamination (see Biochim. Biophs. Acta., 566(2), 259 (1979). However, this deamination process, too, has not been practiced very often due to the reasons noted above.
Due to these difficulties, the isolation and purification of DDA has been practiced only in laboratories by means of repeated liquid chromatography. No commercial process for the purification of DDA is available.
In cases where DDA is produced either microbially or enzymatically from substrates such as 2 ,3,-dideoxyuridine (DDU) or 2,3-dideoxyribose-1-phosphoric acid, the reaction mixture obtained contains, in addition to the desired product (DDA), unreacted substrates, i.e., DDU and adenine ("Ad"), uracil ("U") formed by the decomposition of the substrate, and small quantities of nucleic acids formed as by-products.
Known treatments, such as concentration and recrystallization, are also not suited to obtain highly pure DDA efficiently. This is because both DDU and DDA have a high solubility and therefore could not be separated easily although Ad and U, solubilities of which are relatively small, can be removed by concentration to some extent.
In addition to this, DDA tends to be hydrolyzed under acidic conditions. It is therefore difficult to purify DDA using an ion exchange treatment since an acid is needed for elution of DDA and DDA is consequently hydrolyzed to a 2,3-dideoxyribose residue and an adenine residue.
In view of the advantageous properties of these materials there is thus a strongly felt need for both a more efficient process for their production and for an effective method for purifying the same.